Renal cell carcinoma (RCC) is the most lethal urologic malignancy. The RCC incidence exceeds that of leukemia and pancreatic cancer in men, and equal to that of ovarian and pancreatic cancer in women. The overall age-adjusted RCC incidence is 65.3/100,000 in those >65 years and it now accounts for almost 4% of adult malignancies. Due to increased general diagnostic use of abdominal imaging, the incidental discovery of occult small renal masses has increased. However, radiologic screening is impractical and expensive. Additionally, CT imaging, while detecting renal masses, cannot reliably differentiate RCC from a benign mass. The unmet need is a sensitive and specific tumor marker, for differential diagnosis of imaged renal masses and for population screening and detection of RCC. We have shown that urine aquaporin 1 (AQP1) and perilipin 2 (PLIN2) concentrations are sensitive and specific biomarkers for early noninvasive detection of clear cell or papillary RCC (together accounting for almost 90% of RCC) These biomarkers have cleared the first three hurdles of diagnostic cancer biomarker development: identifying promising biomarkers (phase 1), establishing that the biomarkers identify clinical disease from potential confounding diseases (phase 2), and validating that the biomarkers detect early pre-clinical disease and defined a screen positive rule for urine AQP1 and PLIN2 in a 720 patient cohort (phase 3). We recently developed novel nanotechnology-based assays for AQP1 and PLIN2, achieving greater laboratory sensitivity and specificity, with less expense, and quicker results. Our hypothesis is that nanoAQP1 and nanoPLIN2 assays will have analytical and clinical validity, and clinical utility to achieve rapid differential diagnosis of imaged renal masses in th radiologic setting and for population screening for RCC. We seek to establish phase 4 of biomarker development and define the true detection and the false negative rates of this streamlined nanotechnology-based biomarker assay in a novel prospective 25,000 patient virtual clinical study mimicking population screening, with 1-time cross-sectional evaluation, to measure urine biomarkers, and correlate with clinical diagnoses and available abdominal imaging. The use of nano-AQP1 and nano-PLIN2 assays to differentiate renal tumor subtypes will be tested in another virtual clinical study encompassing 1600 patients with an imaged renal mass. The overall research goal is translation and definitive validation, in a large-scale single-center clinical investigation, of urine AQP1 and PLIN2 as noninvasive biomarkers for 1) differential diagnosis of imaged renal masses, to inform decisions when to surgically intervene, and 2) population screening to detect RCC and enable early and curative intervention. When successfully completed, this research will have translated the basic discovery of RCC biomarkers into a first-ever clinical method for detecting RCC. Thus, our nanotechnology-based biomarker assay to non-invasively diagnose an imaged renal mass and screen for RCC will fundamentally change the course of a silent, lethal disease, and enable better patient outcome.